Differential gene expression from midguts of refractory and susceptible lines of the mosquito, Aedes aegypti, infected with Dengue-2 virus.

Suppressive subtractive hybridization was used to evaluate the differential expression of midgut genes of feral populations of Aedes aegypti (Diptera: Culicidae) from Colombia that are naturally refractory or susceptible to Dengue-2 virus infection. A total of 165 differentially expressed sequence tags (ESTs) were identified in the subtracted libraries. The analysis showed a higher number of differentially expressed genes in the susceptible Ae. aegypti individuals than the refractory mosquitoes. The functional annotation of ESTs revealed a broad response in the susceptible library that included immune molecules, metabolic molecules and transcription factors. In the refractory strain, there was the presence of a trypsin inhibitor gene, which could play a role in the infection. These results serve as a template for more detailed studies aiming to characterize the genetic components of refractoriness, which in turn can be used to devise new approaches to combat transmission of dengue fever.

Identification and characterization of two novel lysozymes from Rhodnius prolixus, a vector of Chagas disease.

Lysozymes have been described in invertebrates as digestive or immune molecules. We report here the characterization of two novel c-type lysozymes, RpLys-A (EU250274) and RpLys-B (EU250275), isolated from the fat body and digestive tract of immune stimulated Rhodnius prolixus, a major vector of Chagas disease. Transcriptional profiles indicate that the temporal and spatial expression patterns of these two peptides are very different. RpLys-A is expressed predominantly in the midgut after ingestion of Trypanosoma cruzi in a bloodmeal, or after injection of bacteria into the hemocoel. RpLys-B is expressed primarily in the fat body after bacterial injection. Phylogenetic alignments indicate that RpLys-A aligns best with molecules from other hemipterans whose major expression is found in the intestinal tract whereas RpLys-B aligns best with mosquito and tick molecules whose expression is found principally in hemocytes and fat body and whose role has been described as immune-related. These data suggest a differential compartmentalized role of two closely related molecules; one for immunity in the hemocoel and the other for digestion in the midgut.

Rhodnius prolixus: identification of immune-related genes up-regulated in response to pathogens and parasites using suppressive subtractive hybridization.

We report the identification of immune-related molecules from the fat body, and intestine of Rhodnius prolixus, an important vector of Chagas disease. Insects were challenged by introducing pathogens or Trypanosoma cruzi, the parasite that causes Chagas disease, into the hemocoel. RNA from intestines, or fat body were isolated 24h after stimulation. We used suppressive subtractive hybridization to identify immune-related genes, generated three subtracted libraries, sequenced the clones and assembled the sequences. The functional annotation revealed expressed sequence tags (ESTs) generated in response to various stimuli in all tissues, and included pathogen recognition molecules, regulatory molecules, and effector molecules.